In Silico Predictions of N-Terminal Residue Cleavage Preferences and Class-Specific 4MDM Modulation in LTA4H

Abstract

Leukotriene A4 hydrolase (LTA4H), a bifunctional zinc metalloenzyme, plays a critical role in human inflammatory pathways. It exclusively initiates inflammation via generation of the pro-inflammatory chemoattractant leukotriene B4. It also creates anti-inflammatory signals by processing endogenous peptides via its aminopeptidase activity. The tripeptide Pro-Gly-Pro (PGP) was long thought to act as a pro-inflammatory chemoattractant whose cleavage by LTA4H resolves inflammation. Recent findings challenge PGP’s physiological role, leaving the key substrate or substrates that mediate the anti-inflammatory activity shrouded in mystery. The small molecule 4MDM enhances LTA4H’s anti-inflammatory AP activity, acting as a pro-survival agent in several mouse studies of inflammation. To narrow the possible N-terminal sequences influenced by 4MDM, we in silico screened all 8,000 canonical tripeptides against LTA4H in the presence or absence of 4MDM. The N-terminal residue appeared to drive predicted binding affinity across the dataset. Bulky aromatic (Trp-X-X, Tyr-X-X) and Arg-X-X tripeptides exhibited enhanced binding in the absence of 4MDM but significantly reduced binding in its presence. 4MDM enhanced binding of Ala-XX tripeptides. Both observations are consistent with biochemically determined cleavage rates. We discuss the implications of how the binding strength varies with each amino acid at the P1 position, including the possibility that new, biologically relevant substrates of LTA4H can be identified.